Visualizing Secretory Cargo Transport in Budding Yeast

摘要

Budding yeast is an excellent model organism for studying the dynamics of the Golgi apparatus. To characterize Golgi function, it is important to visualize secretory cargo as it traverses the secretory pathway. We describe a recently developed approach that generates fluorescent protein aggregates in the lumen of the yeast endoplasmic reticulum and allows the fluorescent cargo to be solu-bilized for transport through the Golgi by addition of a small-molecule ligand. We further describe how to generate a yeast strain expressing the regulatable secretory cargo, and we provide protocols for visualizing the cargo by 4D confocal microscopy and immunoblotting.